On the evolution of lignin biosynthesis in plants

植物木质素生物合成的进化

• 批准号: RGPIN-2019-04562
• 负责人: Ehlting, Juergen
• 金额: $ 3.42万
• 依托单位: University of Victoria
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=689545
• 关键词: evolution; lignin; biosynthesis; plants

Lignin is a defining feature of all vascular plants. This phenolic polymer is incrusted into woody secondary cell walls to provide mechanical stability and hydrophobicity necessary for upright growth and long-distance water transport. Despite common occurrence in all vascular plant lineages, the biosynthetic route to lignin appears to be different in different plant lineages. In flowering plants, the rate limiting step towards the majority of lignin monomers is catalyzed by the enzyme 4-coumaroyl-shikimate-3'-hydroxylase (C3'H / CYP98). However, in lycopods, an early diverging vascular plant lineage, biosynthesis to the same lignin monomers proceeds through a different route. Fittingly, we have recently shown that in lycopods CYP98 is unable to metabolize 4-coumaroyl-shikimate, but utilizes different substrates, presumably to produce defense related secondary metabolites derived from the same pathway. Surprisingly, a fern CYP98 was also unable to use 4-coumaroyl-shikimate, and thus monolignol biosynthesis in ferns also appears to employ distinct intermediates. Even in conifers, CYP98 substrate profiles are distinct from lignin related C3'H enzymes. We thus hypothesize that lignin biosynthesis was modified significantly independently multiple times during land plant evolution. We will test this hypothesis by functionally characterizing the single CYP98s from a conifer and from a fern species. Besides detailing biochemical characterizations, we will need to compare gene expression profiles of these CYP98 genes with genes known to be involved in lignin biosynthesis through analysis of large-scale gene expression data. These will also be mined to identify novel candidate genes involved in lignin biosynthesis in these lineages. Importantly, we need to establish approaches to generate mutants in these non-model species to functionally test a direct involvement of these genes in wood formation. We will generate loss-of-function mutants that will be characterized for changes typically found in flowering plant cyp98 mutants, including changes in lignin composition. Alterations in leaf morphology and metabolite composition will be assessed as these are affected in a non-lignin related moss cyp98 mutant. These experiments will show whether CYP98 is involved in lignin biosynthesis in ferns and conifers or if alternative pathways have evolved. If the latter holds true, candidate genes identified from the gene expression data will be tested for their ability to rescue the cyp98a3 loss of function phenotype in the model plant Arabidopsis and, if warranted, then further characterized biochemically and through reverse genetic approaches. Taken together, this program will test the evolutionary trajectory of lignin biosynthesis and may identify evolutionary optimizations of a biochemical pathway that was crucial for plants to conquer land and that defines wood properties and its uses by humans.

木质素是所有维管植物的定义特征。这种酚类聚合物被镶嵌在木质次生细胞壁中，以提供直立生长和长距离水运输所必需的机械稳定性和疏水性。尽管在所有维管植物谱系中普遍存在，但木质素的生物合成途径在不同的植物谱系中似乎是不同的。在开花植物中，朝向大多数木质素单体的限速步骤由酶4-香豆酰-莽草酸-3 '-羟化酶（C3' H/CYP 98）催化。然而，在石松属植物（一种早期分化的维管植物谱系）中，相同木质素单体的生物合成通过不同的途径进行。我们最近发现，在石松类动物中，CYP 98不能代谢4-香豆酰-莽草酸，但利用不同的底物，可能产生来自相同途径的防御相关次级代谢产物。令人惊讶的是，蕨类植物CYP 98也不能使用4-香豆酰-莽草酸，因此蕨类植物中的单萜醇生物合成似乎也采用了不同的中间体。即使在针叶树中，CYP 98底物谱也不同于木质素相关的C3 'H酶。因此，我们假设木质素的生物合成在陆地植物进化过程中多次独立地进行了显着修改。我们将测试这一假设的功能特性的单一CYP 98从针叶树和蕨类植物物种。除了详细说明生化特征外，我们还需要通过分析大规模基因表达数据来比较这些CYP 98基因与已知参与木质素生物合成的基因的基因表达谱。这些也将被挖掘，以确定新的候选基因参与这些谱系中的木质素生物合成。重要的是，我们需要建立方法来产生突变体在这些非模式物种功能测试这些基因在木材形成的直接参与。我们将产生功能丧失的突变体，其特征在于通常在开花植物cyp 98突变体中发现的变化，包括木质素组成的变化。将评估叶形态和代谢物组成的改变，因为这些在非木质素相关的苔藓cyp 98突变体中受到影响。这些实验将显示CYP 98是否参与蕨类植物和针叶树中的木质素生物合成，或者是否已经进化出替代途径。如果后者成立，将测试从基因表达数据中鉴定的候选基因在模式植物拟南芥中拯救cyp 98 a3功能丧失表型的能力，并且如果有必要，则通过生物化学和反向遗传方法进一步表征。总之，该计划将测试木质素生物合成的进化轨迹，并可能确定生物化学途径的进化优化，该途径对植物征服土地至关重要，并定义了木材特性及其对人类的用途。